A complex cascade of events follows the activation of a cell by a polypeptide hormone, beginning with elevation of cyclic AMP and leading to the phosphorylation of proteins. The proposed research focuses on one component of this cascade, cyclic AMP-dependent protein kinase and the endogenous protein inhibitor of this enzyme. The levels of both the kinase and the inhibitor protein have been observed to change in vivo with changes in levels of the calcium regulating hormones, parathyroid hormone and/or 1,25-dihydroxyvitamin D3. It is proposed to examine in more detail the hormonal regulation of these proteins in chick kidney cell cultures. The inhibitor protein will be purified from chick brain and kideny and monoclonal antibodies prepared against it. These will be used to develop a radioimmunoassay for the inhibitor protein, overcoming present technical difficulties in assaying the protein. In addition, the kinase and the inhibitor protein will be visualized by fluorescence microscopy to determine their cellular and subcellular distribution. The information to be obtained regarding the modulation of these important regulatory proteins and the mechanism by which it occurs will deepen our understanding of the hormonal regulation of cellular function.